Headspace gas chromatography (GC) has been used since the 1980s, but only recently has it become part of mainstream pharmaceutical analysis. The United States Pharmacopeia (USP) incorporated the technique in 2007 into its General Chapter <467> "Residual Solvents." USP defines the use of headspace GC for the analysis of organic volatile impurities (OVIs) or, more commonly, residual solvents (1). A primary benefit of using headspace GC is that one can analyze small amounts of residual solvents buried in a large amount of matrix (i.e., active pharmaceutical ingredient) without having to inject the matrix onto the chromatography column. This technique results in clean, easy sample preparation coupled with less wear on the chromatography columns and the GC instrument because there is less compound passing through the injector or injection liner and detector.

Definition of headspace

The process described in this study is more accurately known as static headspace. However, because no other headspace techniques will be discussed, only the term headspace will be used. The methodology for headspace is quite simple. A sample is diluted and sealed in a closed container that is large enough to allow some headspace in the vial where vapors can collect. The sample is thermostated for a period of time to allow the solvents in the matrix to volatilize, enter the gas phase, and reach equilibrium with the remaining solvent(s) in the vial. This gas phase is the headspace, which is sampled and injected onto the column using pressure as the injection mode. The injection precision of a headspace injector can consistently give < 10.0% relative standard deviation (RSD) and frequently gives ≤ 2.0% RSD, similar to the precision found with liquid-chromatography injection.

The authors studied small, basic organic amines, which are commonly used in pharmaceutical processes instead of more traditional bases such as sodium hydroxide or sodium ethoxide. Use of these amines presents several analytical challenges. The International Conference on Harmonization (ICH) guidelines do not include limits for many such amines; pyridine being the major exception (2). The target value for the specification for such amines can be assigned by the chemist or quality assurance department, which can lead to the same amine having very different set limits. Because specifications are often set at the lowest level the process will tolerate, having an efficient method that can detect low levels of these small organic amines is important. Additional analytical problems with these compounds include non-Gaussian peak shape due to interactions with the GC column, and low flame-ionization detection (FID) based on the size of the compound and low amounts of oxidizable (via burning) carbon groups.

Experimental considerations

This investigation focused on some headspace parameters that should be considered when developing a headspace method, with the goal of maximizing the signal and sensitivity while efficiently using time and materials. All these parameters are important in the early stages of drug development, however, a fast, efficient, sensitive method is also beneficial to the quality-control laboratories that release approved drugs. The authors evaluated experimental parameters including sample of volume in the headspace vial, incubation time in the headspace oven, and the composition of the diluent. The three amines chosen (triethylamine, n-butylamine, and allylamine) represent a cross-section of organic amines that have been analyzed by the authors during scale-up processes.

Figure 1

Sample volume. Headspace sample vials typically come in 10-mL and 20-mL sizes. Often, the analytical chemist needs to analyze a very small amount of a residual solvent in a matrix, which raises the issue of sensitivity. Traditionally, the analytical chemist would increase the concentration of a sample or inject more sample onto a column to get a better signal, thereby increasing the signal strength. With headspace, more sample volume does not always provide the expected increase in area counts because the greater the sample volume, the smaller the actual headspace volume becomes (see Figure 1). The article describes the effect that sample volume has on response for triethylamine, n-butylamine, and allylamine in different diluents.

Incubation time. Solvent-vapor equilibria play a role in headspace analysis. The more readily a solvent can be evaporated into the headspace, the more of that particular solvent will be injected onto the column. Incubation of the headspace vial is an important consideration in headspace-GC method development. If the sample is incubated for too short a period of time, less of the analyte will be in the headspace, which can affect overall area counts. After a certain point, however, the analyte and the solution from which it came will settle into equilibrium; more incubation will not result in any more sample entering the vapor phase and may result in sample degradation or cause secondary reactions. The authors varied the incubation time of triethylamine, n-butylamine, and allylamine samples and reported the change in solvent response over time.

Diluent composition. Salt raises the boiling point of water by affecting the equilibrium of a solvent with its vapor. Because headspace also depends on solvent-vapor equilibria, such techniques can also be used to increase vapor concentration in a headspace-GC sample. Salts have been used in headspace analysis (3–5); however, this technique works best for aqueous-based solvents, which are not the best solvents for GC analysis. The authors, therefore, used high-boiling solvents such as dimethylsulfoxide (DMSO) and dimethylformamide (DMF) as sample diluents. The authors varied the percent of water (aqueous) in the diluent to see whether mixing in a solvent with a higher vapor pressure would affect the concentration of volatile solvents in the headspace.